Protocol for clickable photoaffinity labeling and quantitative chemical proteomics

Lee, Wan-Kyu; Huang, Zhen; Ende, Christopher W. am; Seneviratne, Uthpala

doi:10.1016/j.xpro.2021.100593

Cited by 6 publications

(4 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC) (Ong et al, 2002) introduces isotopes during the cell culture, thereby reducing variations in sample processing by combining individual samples at an earlier stage. Pairwise SILAC experiments can be used to assess probe or small molecule selectivity (Adibekian et al, 2011; Dai et al, 2018; Dettling et al, 2020; Everley et al, 2007; Kamat et al, 2015; Lanning et al, 2014; Lee et al, 2021). Furthermore, a common SILAC cell lysate reacted with a probe can be used as the quantitation reference for untargeted and targeted quantitation in chemical proteomics (S. Li et al, 2017).…”

Section: Chemical Reactions Of the Proteomementioning

confidence: 99%

“…The direct attachment of a reporting moiety to a drug can also alter its selectivity and potency (P. Y. Yang et al, 2011). Therefore, the activity of these probes should be compared to their parent compounds to evaluate any potential effects of the added reporter moiety (Lee et al, 2021; Wijeratne et al, 2018).…”

Section: Proteomics Measurements Utilizing Reactive Reagents With a Tagmentioning

confidence: 99%

“…Probes with highly reactive warheads, such as active esters, may not be the most appropriate for MS analyses, as they can lead to less selective reactions and more complex sample matrixes, even with enrichment strategies. Using parent compounds as competitors in competitive proteomic reactivity profiling can help differentiate nonspecific probe‐protein reactions when determining protein targets of small reactive molecules (Lee et al, 2021; Wijeratne et al, 2018). Achieving the right balance between selectivity and detectability is essential for effectively studying specific protein‐probe reactions using chemical proteomics.…”

Section: Perspectives On Ms Of the Proteomic Reactivity Landscapementioning

confidence: 99%

“…Yang et al, 2011). Therefore, the activity of these probes should be compared to their parent compounds to evaluate any potential effects of the added reporter moiety (Lee et al, 2021;Wijeratne et al, 2018).…”

Section: Selective Compound-centric Probesmentioning

confidence: 99%

See 3 more Smart Citations

Measurement and utilization of the proteomic reactivity by mass spectrometry

Punzalan

Wang

Bajrami

et al. 2023

Mass Spectrometry Reviews

View full text Add to dashboard Cite

Chemical proteomics, which involves studying the covalent modifications of proteins by small molecules, has significantly contributed to our understanding of protein function and has become an essential tool in drug discovery. Mass spectrometry (MS) is the primary method for identifying and quantifying protein‐small molecule adducts. In this review, we discuss various methods for measuring proteomic reactivity using MS and covalent proteomics probes that engage through reactivity‐driven and proximity‐driven mechanisms. We highlight the applications of these methods and probes in live‐cell measurements, drug target identification and validation, and characterizing protein‐small molecule interactions. We conclude the review with current developments and future opportunities in the field, providing our perspectives on analytical considerations for MS‐based analysis of the proteomic reactivity landscape.

show abstract

Section: Chemical Reactions Of the Proteomementioning

confidence: 99%

Section: Proteomics Measurements Utilizing Reactive Reagents With a Tagmentioning

confidence: 99%

Section: Perspectives On Ms Of the Proteomic Reactivity Landscapementioning

confidence: 99%

Section: Selective Compound-centric Probesmentioning

confidence: 99%

See 2 more Smart Citations

Measurement and utilization of the proteomic reactivity by mass spectrometry

Punzalan

Wang

Bajrami

et al. 2023

Mass Spectrometry Reviews

View full text Add to dashboard Cite

show abstract

Proteomic Profiling of Antimalarial Plasmodione Using 3‐Benz(o)ylmenadione Affinity‐Based Probes

Iacobucci,

Monaco,

Hovasse

et al. 2024

ChemBioChem

View full text Add to dashboard Cite

Understanding the mechanisms of drug action in malarial parasites is crucial for the development of new drugs to combat infection and to counteract drug resistance. Proteomics is a widely used approach to study host‐pathogen systems and to identify drug protein targets. Plasmodione is an antiplasmodial early‐lead drug exerting potent activities against young asexual and sexual blood stages in vitro with low toxicity to host cells. To elucidate its molecular mechanisms, an affinity‐based protein profiling (AfBPP) approach was applied to yeast and P. falciparum proteomes. New (pro‐)AfBPP probes based on the 3‐benz(o)yl‐6‐fluoro‐menadione scaffold were synthesized. With optimized conditions of both photoaffinity labeling and click reaction steps, the AfBPP protocol was then applied to a yeast proteome, yielding 11 putative drug‐protein targets. Among these, we found four proteins associated with oxidoreductase activities, the hypothesized type of targets for plasmodione and its metabolites, and other proteins associated with the mitochondria. In Plasmodium parasites, the MS analysis revealed 44 potential plasmodione targets that need to be validated in further studies. Finally, the localization of a 3‐benzyl‐6‐fluoromenadione AfBPP probe was studied in the subcellular structures of the parasite at the trophozoite stage.

show abstract

Investigation of Glycosylphosphatidylinositol (GPI)‐Plasma Membrane Interaction in Live Cells and the Influence of GPI Glycan Structure on the Interaction

Kundu,

Jaiswal,

Babu Mullapudi

et al. 2023

Chemistry A European J

View full text Add to dashboard Cite

Glycosylphosphatidylinositols (GPIs) need to interact with other components in the cell membrane to transduce transmembrane signals. A bifunctional GPI probe was employed for photoaffinity‐based proximity labelling and identification of GPI‐interacting proteins in the cell membrane. This probe contained the entire core structure of GPIs and was functionalized with photoreactive diazirine and clickable alkyne to facilitate its crosslinking with proteins and attachment of an affinity tag. It was disclosed that this probe was more selective than our previously reported probe containing only a part structure of the GPI core for cell membrane incorporation and an improved probe for studying GPI‐cell membrane interaction. Eighty‐eight unique membrane proteins, many of which are related to GPIs/GPI‐anchored proteins, were identified utilizing this probe. The proteomics dataset is a valuable resource for further analyses and data mining to find new GPI‐related proteins and signalling pathways. A comparison of these results with those of our previous probe provided direct evidence for the profound impact of GPI glycan structure on its interaction with the cell membrane.

show abstract

Protocol for clickable photoaffinity labeling and quantitative chemical proteomics

Cited by 6 publications

References 20 publications

Measurement and utilization of the proteomic reactivity by mass spectrometry

Measurement and utilization of the proteomic reactivity by mass spectrometry

Proteomic Profiling of Antimalarial Plasmodione Using 3‐Benz(o)ylmenadione Affinity‐Based Probes

Investigation of Glycosylphosphatidylinositol (GPI)‐Plasma Membrane Interaction in Live Cells and the Influence of GPI Glycan Structure on the Interaction

Contact Info

Product

Resources

About